Objective To observe the expression of GdCl3 on Toll-like receptors (TLRs) of RAW264.7 from murine macrophage cell line induced by lipopolysaccharide (LPS) stimulation. Methods Cells were divided into 3 groups: blank group, LPS group and GdCl3 group. And these cells dyed by goat anti-mouse TLR2/4 poly-antibody and anti-goat IgG labelled with fluorescein isothiocyanate (FITC). The synthesis of TLR2/4 protein were determined by flow cytometry (FCM) analysis and reverse transcription polymerase chain reaction (RT-PCR) analyzed their gene expression. Cell supernatants were taken to measure TNF-α production following the ELISA (enzyme-linked immunosorbent assay) protocol. Results The expressions of TLR2/4 protein and mRNA in GdCl3 group under action of different concentration of GdCl3〔TLR2/4 protein, 200 μmol/L: (70.2±1.28)%/(66.7±2.59)%, 400 μmol/L: (64.9±1.43)%/(60.4±1.25)%, 2 000 μmol/L: (47.4±0.98)%/(32.1±0.74)%; TLR2/4 mRNA (the value of absorbance), 200 μmol/L: (76.42±2.76)/(101.72±3.14), 400 μmol/L: (75.60±3.76)/(89.65±5.17), 2 000 μmol/L: (64.22±4.67)/(78.44±4.88)〕 were significantly lower than those of in LPS group 〔TLR2/4 protein: (94.4±1.76)%/(95.7±0.87)%, P<0.01; TLR2/4 mRNA: (127.64±3.25)/(119.82±5.59), P<0.05, P<0.01〕. The expression of TNF-α in GdCl3 group under action of different concentration of GdCl3〔200 μmol/L: (2 540±77) pg/ml, 400 μmol/L: (2 041±106) pg/ml, 2 000 μmol/L: (1 020±220) pg/ml〕 was also significantly lower that that of in LPS group 〔(4 688±127) pg/ml, P<0.01)〕. Conclusion GdCl3 significantly inhibits TLR expression and secretion of TNF-α under the condition of LPS stimulation in vivo.
目的探讨FasL在重症急性胰腺炎(SAP)大鼠肺泡巨噬细胞(AM)凋亡机理中的作用。 方法SD大鼠按数字表法随机分为对照组、SAP组及氯化钆(GdCl3)组3组,每组16只。SAP模型制成6 h后,经支气管肺泡灌洗获取AM。取右肺下叶行HE染色检查,透射电镜观察和双染色法检测AM凋亡情况,用蛋白免疫印迹法(Western blot法)检测各组AM中FasL蛋白表达水平。 结果GdCl3组电镜下可见AM典型凋亡形态学特征,AM凋亡率为(22.48±1.44)%,明显高于对照组〔(11.28±1.01)%〕及SAP组〔(6.86±1.35)%〕,其差异有统计学意义(P<0.05);GdCl3组FasL蛋白相对表达量为(1.230±0.041)%,较对照组〔(0.936±0.024)%〕和SAP组〔(0.704±0.011)%〕明显增高(P<0.05)。AM凋亡率与AM中FasL蛋白表达水平呈线性正相关关系(R2=0.766,P<0.01)。 结论GdCl3可能通过激活FasL蛋白表达诱导SAP大鼠AM发生凋亡。